Go with the flow: Better SERS detection with confined streams

Ezine

Published: Oct 21, 2013

Author: Jon Evans

Channels: Detectors

Silver and gold

Surface-enhanced Raman spectroscopy (SERS) is a detection technique sensitive enough to detect single molecules. Like conventional Raman spectroscopy, SERS detects analytes based on the way they scatter a beam of light from a laser, as individual analytes tend to scatter light in characteristic ways. In SERS, however, this effect is greatly enhanced by conducting it on a surface covered with tiny gold or silver nanoparticles, making it sensitive enough to detect individual molecules.

This kind of sensitivity, together with the fact that gold and silver nanoparticles can be deposited on almost any surface, means that SERS should make an ideal detection technique for microchip versions of liquid chromatography (LC) and capillary electrophoresis (CE). The idea being to deposit the nanoparticles at the far end of a separation capillary or channel, and then detect the separated analytes as they flow through.

Should, though, is the operative word, because when scientists such as Zachary Schultz at the University of Notre Dame in Indiana, US, have tried combining SERS with LC and CE, they’ve found it doesn’t work as well as they’d hoped. ‘My research group has been using nanostructures to do enhanced Raman spectroscopy but were always perplexed by the low efficiency of detection in solution,’ Schultz told separationsNOW. ‘We had results showing sensitivity was not the problem, but rather keeping molecules on the surface.’

Twin streams

To be detected by SERS, analytes have to be fairly close to the nanoparticles, within around 10nm, but obviously the majority of the analytes flowing through the capillary or channel aren’t that close to the nanoparticles and thus can’t be detected by SERS. This means that the analyte concentration needs to be quite high, to ensure that at least some analytes are close to the nanoparticles, negating the technique’s high sensitivity.

Several research groups have come up with ways to surmount this problem, which mainly involve getting the analytes to stick to the nanoparticles, such as by making the analytes magnetic or chemically modifying them so that they bind with the nanoparticles. But these approaches add extra complexity to the analytical process. Quite by chance, Schultz recently stumbled on an alternative and much simpler answer.

‘I was listening to a seminar by one of my co-authors, Oluwatosin Dada, about work he was doing with sheath-flow fluorescence detection and it occurred to me that the same effect could be used to confine molecules near a SERS substrate,’ Schultz explains. This detection method utilizes a technique known as hydrodynamic focusing, in which a slow-moving analyte stream is introduced into a much faster moving sheath stream. As the fast-moving sheath stream rushes past, it creates a physical barrier that confines the analyte stream into a narrow flow. Hydrodynamic focusing is regularly used to confine cells in flow cytometry and Schultz realized it could also confine analytes to the bottom of a capillary or a channel, keeping them close to where the nanoparticles are deposited.

Inside the chamber

Together with Dada and two other colleagues, Schultz constructed an analytical system comprising a thin capillary inside a larger chamber. The fast sheath flow is pumped through the chamber, confining analytes emerging from the capillary into a narrow flow at the bottom of the chamber, where Schultz deposits a mixture of gold and silver nanoparticles.

Trying this system out on a rhodamine dye, Schultz found the dye could be detected by SERS at concentrations of just 1nM, three orders of magnitude better than could be achieved without hydrodynamic focusing. Schultz and his colleagues have since gone on to show that this system works with CE and will soon test it with LC.

‘We have already coupled the detector to CE separations,’ he says. ‘We have done a variety of dye molecules, including isobaric rhodamine dyes, but more recently we have begun looking at amino acids as a model system with intrinsic chemical variability.’

One of the reasons Schultz is particularly excited about this novel SERS detection system is because it can potentially provide chemical information about analytes that other techniques can’t match. ‘The applications we hope to advance are small molecules that don't ionize well or are isobaric and [so] are difficult to study by mass spectrometry,’ he says.